Auxin Response Factors as a model of transcriptional control

生长素反应因子作为转录控制模型

• 批准号: 10188569
• 负责人: Lucia Strader
• 金额: $ 39.13万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10188569
• 关键词: 4-ethoxymethylene-2-phenyl-2-oxazoline-5-one; Automobile Driving; Auxins; Back; Behavior; Binding; Biochemistry; Biophysics; Charge; Chromatin Remodeling Factor; Complex; Crystallization; Cullin Proteins; Development; Disease; Electrostatics; Event; F Box Domain; F-Box Proteins; Face; Family; Gene Activation; Genetic; Genetic Transcription; Hormones; Knowledge; Ligands; Mediating; Mission; Modeling; Molecular; Nature; Organism; Phase Transition; Physical condensation; Plants; Point Mutation; Positioning Attribute; Process; Proteins; Reagent; Regulation; Repression; Repressor Proteins; Research; Side; Signal Transduction; Structure; System; Transcription Coactivator; Transcription Repressor; Transcriptional Regulation; United States National Institutes of Health; derepression; gene repression; insight; interdisciplinary approach; member; plant growth/development; protein degradation; protein folding; receptor; recruit; response; transcription factor; transport inhibitor; ubiquitin-protein ligase

Project Summary Hormone-mediated modulation of gene activation or repression through transcription factors is central to all organisms. AUXIN RESPONSE FACTOR (ARF) transcription factors are critical modulators of plant growth and provide an ideal model for exploring hormone control of gene activation and repression. Of the 23-member ARF family, five are considered transcriptional activators and 18 are considered transcriptional repressors, allowing for study of both of these activities in a single family. Under low auxin concentrations, Aux/IAA proteins repress ARF transcription factors via direct interaction and recruitment of chromatin remodeling factors. When auxin concentrations are high, a co-receptor complex, comprised of an F-box protein from the TRANSPORT INHIBITOR REPONSE1 (TIR1) family and an Aux/IAA repressor protein, directly binds auxin. The F-box protein participates in a Skp1-Cullin-F-box (SCF) E3 ubiquitin ligase, which targets the Aux/IAA protein for degradation. This degradation event relieves ARF transcription factor repression, allowing auxin-regulated transcription. This receptor-ligand interaction allows a very short signal transduction chain to facilitate rapid transcriptional responses to auxin. To understand the molecular underpinnings of ARF-ARF and ARF-Aux/IAA interactions, our lab solved the structure of the domain driving these interactions, finding that it folds into a Type I/II Phox and Bem1 (PB1) domain. Within this domain, there is a positively charged and a negatively charged electrostatic face on opposing sides, creating a Janus-like protein fold. This allows for front-to-back ARF oligomerization (similar to a set of bar magnets) in the packed crystal, in solution, and in the plant. In addition to the well-studied repression – derepression mechanism of regulation, our lab has discovered that activity of a subset of ARFs can be regulated by protein phase transition driven by the combination of PB1 oligomerization and an intrinsically disordered region. Phase transition of these ARFs appears to modulate responsiveness to auxin in a developmentally relevant context. We have further found that many ARFs are regulated by proteasomal degradation and have identified an E3 ubiquitin ligase involved in this process. Finally, ARF interactions can be easily manipulated using PB1 domain point mutations, allowing us to direct ARF interactions for study. Using ARFs as a model will allow us to interrogate transcription factor function in an easily manipulated system to yield broad insight into many transcription factors. We are aided in our efforts by our multidisciplinary approach, extensive auxin-related molecular toolkit, and unique reagents generated by our lab. Our lab's expertise in genetics and biochemistry/biophysics, combined with our recent discoveries of ARF condensation and proteasomal degradation, makes us well positioned to drive forward our understanding of phase transition and other mechanisms in regulation of transcription factor activity.

项目摘要 激素介导的调节基因激活或抑制通过转录因子是核心的所有 有机体生长素反应因子（ARF）转录因子是植物生长的重要调节因子 为探索激素对基因激活和阻遏的调控提供了理想的模型。在23名成员中， ARF家族中，5个被认为是转录激活因子，18个被认为是转录抑制因子， 允许在一个家庭中研究这两种活动。 在低生长素浓度下，Aux/IAA蛋白通过直接相互作用抑制ARF转录因子 和染色质重塑因子的募集。当生长素浓度高时，一种辅助受体复合物， 由来自转运抑制剂应答1（TIR 1）家族的F-box蛋白和Aux/IAA蛋白组成， 阻遏蛋白，直接结合生长素。F-box蛋白参与Skp 1-Cullin-F-box（SCF）E3泛素化 连接酶，其靶向Aux/IAA蛋白进行降解。这种降解事件缓解了ARF转录 因子抑制，允许生长素调节的转录。这种受体-配体相互作用允许非常短的 信号转导链，以促进生长素的快速转录反应。 为了了解ARF-ARF和ARF-Aux/IAA相互作用的分子基础，我们的实验室解决了 驱动这些相互作用的结构域的结构，发现它折叠成I/II型Phox和Bem 1（PB 1） 域在这个区域内，有一个带正电荷和一个带负电荷的静电面， 形成一个类似两面神的蛋白质折叠这允许前后ARF寡聚化（类似于 一组条形磁体）在堆积晶体中、在溶液中和在工厂中。 除了研究充分的抑制-去抑制调节机制外，我们的实验室还发现， ARF的一个子集的活性可以通过由PB 1结合驱动的蛋白质相变来调节， 低聚和固有无序区域。这些ARF的相变似乎调节了 在发育相关的背景下对生长素的反应。我们进一步发现，许多ARF是 通过蛋白酶体降解调节，并确定了E3泛素连接酶参与这一过程。 最后，ARF相互作用可以很容易地使用PB 1结构域点突变操纵，使我们能够直接 研究的ARF相互作用。使用ARF作为模型将使我们能够询问转录因子的功能， 易于操作的系统，以产生广泛的洞察到许多转录因子。 我们的多学科方法，广泛的生长素相关分子工具包， 我们实验室生产的独特试剂我们实验室在遗传学和生物化学/生物物理学方面的专业知识， 随着我们最近发现的ARF凝聚和蛋白酶体降解，使我们能够很好地定位， 推动我们对转录因子调控中的相变和其他机制的理解 活动